Producing method of stress reliever

ABSTRACT

Provided is a method of producing a stress reliever that reduces a decrease in productivity of livestock at the time of stress load by feeding the stress reliever to the livestock. The method of producing the stress reliever includes solid-culturing a substrate with the filamentous bacteria, in which the stress reliever reduces a decrease in productivity of livestock at the time of stress load.

FIELD

The present invention relates to a method of producing a stressreliever.

BACKGROUND

Patent Literature 1 cited below has described a method for selectingspecific Aspergillus Oryzae (IK-05074 strain) that excellently breeds ina culture medium to which sodium deoxycholate is added and inoculatingthe selected Aspergillus Oryzae (IK-05074 strain) to brown rice toproduce a solid-cultured product. In this solid-cultured product,activity of acid-resistant α-amylase, acidic protease, and acidiccarboxypeptidase is higher than that of a cultured product obtained byculturing Aspergillus Kawachii and this solid-cultured product is saidto have the effect of an increase in the body weight of chicks.

CITATION LIST Patent Literature

Patent Literature 1: Japanese Patent Application Laid-open No.2007-325580

SUMMARY Technical Problem

Patent Literature 1 has not described that a decrease in productivity oflivestock is reduced by relieving stress using the solid-culturedproduct of filamentous bacteria when the stress is applied to thelivestock.

When the stress is applied to livestock, the productivity of thelivestock is reduced due to reduction in the ingestion amount offeedstuff or a decrease in the body weight of the livestock.

An object of the present disclosure is to provide a method of producinga stress reliever that reduces the decrease in productivity of livestockat the time of stress load by orally feeding the stress reliever to thelivestock.

Solution to Problem

The above-described problem is solved by a method of producing a stressreliever, the method including: solid-culturing a substrate withfilamentous bacteria to obtain a solid-cultured product, wherein thestress reliever reduces a decrease in productivity of livestock at thetime of stress load.

The stress reliever produced by the method of producing the stressreliever preferably reduces one or more decreases in productivity oflivestock at the time of stress load by relieving the stress of thelivestock ingesting the stress reliever, the one or more decreases beingamong body weight reduction, a decrease in the ingestion amount offeedstuff, a decrease in an egg weight, a decrease in an eggshellthickness, and a decrease in a yolk weight.

In the method of producing the stress reliever, the filamentous bacteriaare preferably bacteria not producing mold poison. In the method ofproducing the stress reliever, the bacteria not producing mold poisonare preferably Aspergillus oryzae, Aspergillus sojae, Aspergillusluchuensis, Aspergillus niger, or Aspergillus awamori not producing moldpoison.

In the method of producing the stress reliever, the solid-culturedproduct preferably includes polysaccharides constituting hyphae of thefilamentous bacteria. In the method of producing the stress reliever,the solid-cultured product preferably includes an enzyme havingactivity. In the method of producing the stress reliever, thesold-cultured product preferably includes viable bacteria of thefilamentous bacteria.

In the method of producing the stress reliever, the method may furtherinclude determining the solid-cultured product alone obtained bysolid-culturing the substrate with the filamentous bacteria to be thestress reliever or blending the solid-cultured product obtained bysolid-culturing the substrate with the filamentous bacteria with afeeding substance.

Advantageous Effects of Invention

According to the present disclosure, a method of producing a stressreliever that reduces a decrease in productivity of livestock at thetime of stress load by orally feeding the stress reliever to thelivestock can be provided.

BRIEF DESCRIPTION OF DRAWINGS

FIG. 1 is a graph illustrating the amount of body weight change (g) onthe next day (the first day) after hot environment treatment started.

FIG. 2 is a graph illustrating the amount of body weight change (g) in aperiod from the day (the zeroth day) when the hot environment treatmentstarted to the seventh day.

FIG. 3 is a graph illustrating a relation between the number of daysafter the hot environment treatment started and the ingestion amount (g)of feedstuff.

FIG. 4 is a graph illustrating a relation between the number of daysafter the hot environment treatment started and an egg weight (g).

FIG. 5 is a graph illustrating a relation between the number of daysafter the hot environment treatment started and an eggshell weight (g).

FIG. 6 is a graph illustrating a relation between the number of daysafter the hot environment treatment started and a yolk weight (g).

DESCRIPTION OF EMBODIMENTS

Hereinafter, the embodiment of the method of producing the stressreliever and the stress reliever produced by the method will bedescribed.

The stress reliever is used by orally feeding the stress reliever tolivestock and includes the solid-cultured product of filamentousbacteria. The effect of reducing the decrease in productivity oflivestock can be obtained by orally feeding the stress reliever tolivestock. The stress reliever can be produced by the method ofproducing the stress reliever including solid-culturing a substrate withfilamentous bacteria to obtain the solid-cultured product.

As the filamentous bacteria used in the above production method, thebacteria not producing mold poison are preferably used. Ingestion of thebacteria not producing mold poison to livestock is safe. Examples of thefilamentous bacteria not producing mold poison include Aspergillusoryzae, Aspergillus sojae, Aspergillus luchuensis, Aspergillus niger, orAspergillus awamori not producing mold poison. As these filamentousbacteria, seed bacteria for the fermentation of fermented foods arecommercially available or the filamentous bacteria can be obtained atNITE Biological Resource Center (NBRC). The filamentous bacteria may bea wild strain of which gene is not modified or may be filamentousbacteria of which gene is modified by a gene technological method asdescribed below.

Examples of the mold poison include aflatoxin, deoxynivalenol,ochratoxin, fumonisin, zearalenone, patulin, sterigmatocystin, orfusarium toxin.

The solid-cultured product of the filamentous bacteria preferablyincludes polysaccharides constituting hyphae of the filamentousbacteria. When the cultured product including the polysaccharides andthe filamentous bacteria are fed together to livestock, the decrease inproductivity of livestock caused by stress can be more remarkablyreduced. Although the detailed mechanism is unclear, it is presumed thatthe immunity of the livestock ingesting the polysaccharides is improved,whereby the decrease in productivity of livestock caused by stress isreduced.

The solid-cultured product of the filamentous bacteria preferablyincludes an enzyme having activity. When the enzyme having the activityis, for example, an enzyme that improves the digestion ratio of thefeedstuff, nutrient contents are easily taken into the body of thelivestock by degrading the feedstuff fed by livestock with the enzymeinside the body of livestock and thus feedstuff efficiency can beimproved. This degradation action is performed inside the body oflivestock and thus digestion of the feedstuff by the livestock can bepromoted.

Although the detailed mechanism is unclear, it is presumed that thedecrease in productivity of the livestock caused by stress is reduced byeasily taking the various nutrient contents into the body of thelivestock. In the outside of the body of livestock, the degradationaction can also be obtained in the case where the feedstuff and thesolid-cultured product of the filamentous bacteria are mixed and cultureor enzyme reaction is performed.

The solid-cultured product of the filamentous bacteria preferablyincludes viable bacteria of the filamentous bacteria. For example, whenthe solid-cultured product includes the viable bactera, the productionof the stress reliever can be easily increased by, for example, addingthe substrate described below to the cultured product to performsecondary culture. According to this method, the stress reliever can beremarkably easily obtained because the seed bacteria are not required tobe subdivided and obtained or the seed bacteria are not required to bepurchased.

In order to form the solid-cultured product having the enzyme activity,the solid-cultured product of the filamentous bacteria may be determinedto be the stress reliever without applying excessive heating to thesolid-cultured product. The excessive heating refers to heating to theextent that the enzyme activity is lost. In order that thesolid-cultured product includes the viable bacteria of the filamentousbacteria, the solid-cultured product of the filamentous bacteria may beused as the stress reliever without performing excessive heating to theextent that the filamentous bacteria become extinct.

For example, in the case where the enzyme is cellulase, pectinase, andthe like, these enzymes catalyze the reaction in which cellulose,pectin, and the like included in the feedstuff or the like are degraded.Polysaccharides such as cellulose and pectin are a kind of the componentconstituting the cell walls of plants. Various kinds of thepolysaccharides constituting the cell walls of plants have been known.The forms thereof are various and the constitutions are complex. Inorder to efficiently degrade the cell wall polysaccharide having complexstructures, the degrading enzymes preferably act stepwise. For example,degradation efficiency of the cell walls of plant raw materials includedin the feedstuff is improved by degrading cellulose, pectin, and thelike with the enzymes such as cellulase and pectinase. Consequently, thefeedstuff becomes easily digested.

For example, in the case where the enzyme is tannase, tannase catalyzesthe chemical reaction in which tannin included in the feedstuff or thelike is degraded. Some types of tannin form complexes by stronglybonding to polymers such as proteins. Tannin may exist in the state ofbeing intricately entangled with the components constituting the cellwalls of plants and may inhibit degradation of the cell walls.Degradation of tannin with tannase improves the degradation efficiencyof the cell walls of plant raw materials included in the feedstuff andthus the feedstuff becomes easily digested.

For example, in the case where the enzyme is phytase, phytase catalyzesthe chemical reaction in which inorganic form phosphoric acid isseparated from phytic acid included in the feedstuff and the like. It issaid that phytic acid inhibits absorption of minerals such as calciumand zinc included in the feedstuff into the body of an animal ingestingthe feedstuff. Therefore, degradation of phytic acid with phytaseimproves the absorption ratio of minerals. Phosphorous generated bydegradation of phytic acid is also absorbed into the body of the animalsingesting the feedstuff.

Examples of the enzyme include enzymes selected from the groupconsisting of amylase, alkaline protease, acidic protease, neutralprotease, xylanase, β-glucanase, cellulase, tannase, phytase, lactase,lipase, pectinase such as polygalac-turonase, a xylanase-pectinasecomplex enzyme, and a cellulase-protease-pectinase complex enzyme. Anyof these enzymes are enzymes coded from the genomic DNA of thefilamentous bacteria and expressed from the filamentous bacteria of thewild strain. Gene manipulation may be performed so that two or moreenzymes among these enzymes are highly expressed in the filamentousbacteria.

Genetic transformation may be performed so that the above enzyme ishighly expressed in the filamentous bacteria as compared to thefilamentous bacteria of the wild strain not subjected to genetictransformation by using a known gene technological method. In thefilamentous bacteria, for example, it is known that the promoter ofamylase (AmyB) or the promoter of enolase (enoA) has high expressionamounts. A chimeric gene is obtained by bonding a gene that codes thetarget enzyme and a terminator sequence corresponding to the promotor byusing a known method to these promoters for high expression. When thischimeric gene is introduced to the filamentous bacteria by a knownmethod, the filamentous bacteria in which the target enzyme is highlyexpressed can be obtained. The analysis of the genome sequences of somefilamentous bacteria has already finished and the sequences arepublished in a database. The sequences of the gene that codes the highexpression promoter, the terminator, and the target enzyme are searchedby using such a database and the primer is designed. The designed primerand a template such as cDNA and genomic DNA are used to amplify thedesired gene sequence with PCR. The amplified gene sequence is used whenconducting the genetic transformation. At the time of the genetictransformation, the above chimeric gene may be introduced at the targetposition of the genome by using a known qenome editing method or theabove chimeric gene may be introduced at any position of the genome byintroducing the chimeric gene into the cells of the filamentousbacteria. In order to selectively culture the genetically transformedfilamentous bacteria, known marker genes such as niaD and ptrA may beused.

At the time of cloning the highly expressing gene, the genomic DNA orcDNA of the filamentous bacteria of the same species as the geneticallytransformed filamentous bacteria is preferably used as the template.When the gene incorporated into the genetically transformed filamentousbacteria is derived from the filamentous bacteria belonging to the samespecies, safety of filamentous bacteria is secured because a foreigngene is not incorporated. Such a cloning method is referred to asself-cloning. The method of the self-cloning involves, for example,cloning the desired gene using the genomic DNA of the wild strain of thekoji mold (Aspergillus oryzae, RIB40) obtainable at NBRC (NationalInstitute of Technology and Evaluation) as the template, andsubsequently introducing the cloned gene to a commercially availablekoji mold for sake brewing (Aspergillus oryzae, AOK11).

The solid-cultured product of the filamentous bacteria is notparticularly limited and can be obtained by, for example, the followingmethod. The solid substrate described below is subjected to steaming andcooled. Seed bacteria are inoculated to the cooled substrate. Theinoculated substrate is placed on a culture bed in a culture apparatusand the culture is performed so that the filamentous bacteria arepropagated in the solid culture substrate by passing air in whichtemperature and humidity are controlled through between. the grains ofthe culture substrate. The temperature of the air is not particularlylimited. For example, the temperature is controlled in a range of 20° C.to 45° C. The humidity of the air is not particularly limited. Forexample, the humidity is controlled in a range of 50% to 99% in relativehumidity. By such a method, the solid-cultured product in which thehyphae of the filamentous bacteria are propagated on the surface andinside of the solid-like substrate can be obtained.

In the solid-cultured product in which the hyphae of the filamentousbacteria are propagated on the surface and inside of the solid-likesubstrate, the filamentous bacteria are vigorously propagated using thesubstrate as a nutrition source and thus a large amount of the enzymeshaving activity and a large amount of the polysaccharides constitutingthe hyphae are included. Therefore, according to the solid-culturedproduct, the decrease in productivity of livestock caused by stress canbe remarkably reduced as compared to the case where the spores of thefilamentous bacteria are fed to livestock or the case where the sporesof the filamentous bacteria are mixed with the feedstuff for livestockand the mixed product is orally fed.

When the filamentous bacteria are cultured in a state of solid, morekinds of enzymes are produced as compared to the case where thefilamentous bacteria are cultured in a state of liquid and theproduction amount of individual enzyme also becomes larger. Therefore,when the stress reliever produced by subjecting the filamentous bacteriato a step of culturing the filamentous bacteria in the state of solid isfed to livestock. The effect of reducing the decrease in productivity oflivestock caused by stress becomes larger as compared to the case wherethe product made by culturing the filamentous bacteria in the state ofliquid is fed to livestock. From the above reason, the filamentousbacteria are preferably subjected to the step of culturing thefilamentous bacteria in the state of solid at the time of the productionof the stress reliever.

The above substrate may be, for example, a solid organic substancesuitable for the filamentous bacteria to breed. The solid includes, inaddition to a solid content having hardness, a slurry-like substance ora power grain product. Examples of the substrate include one or moreorganic substances selected from the group consisting of cereals such asbarleycorn, wheat, the bran of wheat, rice, beans, and corn; residues ofprocessed food such as beet pulp, the squeezed lees of oil, and thesqueezed lees of fermented foods; and food residues such as leftoverfoods Examples of the squeezed lees of oil include the squeezed lees ofsoybean, the squeezed lees of rapeseed, the squeezed lees of sesame, andthe squeezed lees of corn.

The dosage form of the stress reliever is not particularly limited. Forexample, the solid-cultured product obtained by solid-culturing thesubstrate with the filamentous bacteria may be determined to be thestress reliever as it is, the solid-cultured product may be pulverizedto form powder, or the solid-cultured product may be determined to be aliquid form or a slurry form by blending a liquid component to thesolid-cultured product when the feedstuff fed to livestock is the liquidform or the slurry form. A feeding substances may be mixed with thesolid-cultured product. Examples of the feeding substance include a bulkincreasing material that enhances mixing ability with feedstuff,additives such as vitamin compounds, products generally fed such asknown feedstuff for livestock, or a mixture of two or more of them. Thesolid-cultured product alone may be determined to be the stressreliever. In the case where the solid-cultured product alone isdetermined to be the stress reliever, for example, the feeding substanceis not added to the solid-cultured product and the solid-culturedproduct is, for example, inspected and packaged to produce a finishedproduct.

The blend ratio of the solid-cultured product and the feeding substanceis not particularly limited. However, blend of an excessive amount ofthe solid-cultured product exhibits tendency in which the effect forrelieving stress is saturated and thus increases the cost. Due to suchtendency, the blend ratio (%) of the solid-cultured product is, forexample, preferably 0.05% by mass to 5.0% by mass, more preferably 0.05%by mass to 4.0% by mass, and further preferably 0.05% by mass to 3.0% bymass. The effect for reducing the decrease in productivity of livestockat the time of stress load can be obtained even when the solid-culturedproduct is mixed with the feeding substance. The effect for relievingstress can be obtained even when the blend ratio of the solid-culturedproduct is, for example, 0.05% by mass, which is a small blend ratio.

Common feedstuffs of livestock are solid in many cases. In the casewhere the liquid cultured product of the filamentous bacteria is mixedwith the solid feedstuff, liquid part of the feedstuff is increased.This may cause deterioration in a taste property for livestock that isused to the solid feedstuff. In the case where the livestock is used tothe solid feedstuff, the dosage form of the stress reliever ispreferably solid. By doing this, deterioration in the taste property forthe livestock can be prevented. When the solid-cultured product isdetermined to be the stress reliever as it is, processing is notnecessary, which is more preferable.

Examples of relieved stress includes the followings: stress by breedinglivestock in a dense state, stress by vaccination, stress by capturinglivestock by hand, stress by restraining livestock, stress at the timeof transporting livestock, stress in slaughterhouses, stress during anincubation period of eggs, stress at the time of cutting beaks or horns,stress by forced molting, stress by early weaning, stress by castration,stress by childbirth, stress by fasting, stress by feeding restriction,stress by placing livestock in a hot environment, stress by placinglivestock in a cold environment, stress by noise, stress by light beams,or stress by weather.

The target livestock to which the stress reliever is fed is notparticularly limited. Examples the livestock include chickens, cows,pigs, horses, donkeys, camels, goats, sheep, or fish such as carp andrainbow trout. The livestock include livestock of which meat is used,livestock of which eggs are used, and livestock of which milk, leathers,furs, hairs, or feathers are used.

The stress reliever has the effect for relieving stress of livestock andreducing the decrease in productivity. The decrease in productivityrefers to the decrease in the production amount of products to be usedsuch as meat, eggs, milk, hairs, and leathers obtained from livestockand deterioration in the quality of the products to be used. Thedecrease in the production amount includes a decrease in the ingestionamount of feedstuff. Examples of the decrease in productivity includebody weight reduction, a decrease in the ingestion amount of feedstuff,a decrease in an egg weight, a decrease in an eggshell thickness, adecrease in a yolk weight, a decrease in a milking amount, hair loss,and deterioration in the quality of the products to be used.

EXAMPLES

Hereinafter, the method of producing the stress reliever and the stressreliever produced by the method will be described with reference toExamples. Examples to be described below are mere limited examples. Thetechnical scope of the present invention is not limited to Examples.

Example 1

To the bran of wheat (wheat bran), water was added and the resultantmixture was stirred, and thereafter the bran of wheat was subjected tosteaming treatment under a condition of 0.2 MPa. The steamed bran ofwheat was cooled down to around 30° C. A certain amount of the seedbacteria of Aspergillus oryzae (AOK11) that is commercially available asa koji mold for sake brewing and does not produce mold poison wasinoculated and the resultant mixture was mixed so as to be uniform. Thewater content of the bran of wheat at the time of inoculation was set to60%. The culture was started after this raw material was accumulated onthe culture bed in the culture apparatus and smoothed so that thethickness of the accumulated raw material was constant. During theculture, air in which temperature and humidity were controlled was fedto the accumulated raw material and the fed air was passed throughbetween the grains of the raw material. At this time, the temperature ofthe fed air was controlled to 25° C. to 40° C. and the relative humidityof the fed air was controlled to 90% to 96% so that the substancetemperature of the raw material is in a range of 30° C. to 38° C. Duringthe culture, the raw material was stirred using a stirring unit equippedwith the culture apparatus. The culture was performed until hyphae ofthe filamentous bacteria covered the surface of the grains of the branof wheat. This solid-cultured product was determined to be the stressreliever according to Example 1 without sterilizing the bacteria byheating or the like in a state where the solid-cultured product wasalone as it was. This stress reliever includes the viable bacteria ofAspergillus oryzae (AOK11) and includes useful enzymes of Aspergillusoryzae (AOK11) retaining enzyme activity.

Example 2

The cultured product of the solid bran of wheat was obtained in the samemanner as the manner in Example 1 except that Aspergillus oryzae (AOK11)that highly expressed tannase (tanA), pectin lyase (pelA) that is a kindof pectinase, phytase (phyA), and polygalac-turonase (pgaB) that is akind of pectinase was used as the seed bacteria. This solid-culturedproduct was determined to be the stress reliever according to Example 2without sterilizing the bacteria by heating or the like in a state wherethe solid-cultured product was alone as it was. Similar to Example 1,this stress reliever includes the viable bacteria and retains enzymeactivity.

The target genes of tannase (tanA), pectin lyase (pelA), phytase (phyA),or polygalac-turonase (pgaB) were incorporated to Aspergillus oryzae(AOK11) by known methods to perform the genetic transformation. At thetime of cloning the target genes, the genomic DNA of Aspergillus oryzae(RIB40) was used as the template. Each of the target genes describedabove was incorporated between the amylase promoter (AmyB promoter)sequence and the amylase terminator (AmyB terminator) sequence that arehigh expression promoters. The expression cassette that was a chimericgene of thus prepared promotor sequence, the target gene sequence, andthe terminator sequence was not incorporated to the plasmid but wasintroduced to Aspergillus oryzae (AOK11) by a chloroplast-PEG method.These gene sequences can be searched using a koji mold genome database(www.aspgd.org/) and the database of glycosyl hydrolase CAZy(www.cazy.org/fam/acc_GH.html).

[Observation of Body Weight Change]

A plurality of 14-day old hens for meat (Chunky) were bred by dividinginto Experimental Section 1 to Experimental Section 3 described below toobserve an effect in which hot environment stress affected the bodyweight change of the hens.

[Experimental Section 1]

As the feedstuff for the hens, from 14 days old to 20 days old, thestress reliever in Example 2 was mixed with the commercially availablefeedstuff for chick breeding (blended feedstuff for broiler fattening informer period Power Chicken, manufactured by NICHIWA SANGYO CO., LTD.)so that the stress reliever was contained in 2.0% by mass and theresultant mixture was fed to the hens. After 21 days old, the stressreliever in Example 2 was mixed with the feedstuff for latter period(Daisu-Meijin, manufactured by CHUBUSHIRYO CO., LTD.) so that the stressreliever was contained in 2.0% by mass and the resultant mixture was fedto the hens. On the day when the hens became 35 days old (the zeroth daydescribed below), the hens were transferred to a poultry house having ahot environment to breed the hens. The hot environment refers tobreeding in a poultry house having no windows where temperature controlwith an air conditioner is not performed. The temperature was in a rangeof 26° C. to 32° C. Experimental Section 1 included four hens.

[Experimental Section 2]

The hens were bred in the hot environment in the same manner as themanner in Experimental Section 1 except that the feedstuff was changedso that the stress reliever in Example 1 was contained in the sameformer period feedstuff or latter period feedstuff used in ExperimentalSection 1 in 2.0% by mass. Experimental Section 2 included six hens.

[Experimental Section 3]

The hens were bred in the hot environment in the mane manner as theExperimental Section 1 except that the feedstuff was changed so that thebran of wheat not subjected to the solid culture with the filamentousbacteria was contained in the same former period feedstuff or latterperiod feedstuff used in Experimental Section 1 in 2.0% by mass.Experimental Section 3 included five hens.

[Experimental Section 4]

As the feedstuff for the hens, from 14 days old to 20 days old, the branof wheat not subjected to the solid culture with the filamentousbacteria was mixed with the commercially available feedstuff for chickbreeding (blended feedstuff for broiler fattening in former period PowerChicken, manufactured by NICHIWA SANGYO CO., LTD.) so that the bran ofwheat was contained in 2.0% by mass and the resultant mixture was fed tothe hens. After 21 days old, the bran of wheat not subjected to thesolid culture with the filamentous bacteria was mixed with the feedstufffor latter period (Daisu-Meijin, manufactured by CHUBUSHIRYO CO., LTD.)so that the bran of wheat was contained in 2.0% by mass and theresultant mixture was fed to the hens. On the day when the hens became35 days old (zeroth day), the hens were transferred to a poultry househaving a thermal neutral zone to breed the hens. The thermal neutralzone refers to a temperature range suitable for homoiothermic animals.In this experiment, the air temperature was controlled to 22° C., whichwas suitable temperature to breed hens. Experimental Section 4 includedsix hens.

In Experimental Sections 1 to 4, the day when the hens were transferredto the hot environment or the thermal neutral zone to start breeding isdetermined to be the zeroth day. The next day is described as the firstday, the day after the next day is described as the second day, and soforth.

The body weight (g) on the zeroth day was subtracted from the bodyweight (g) on the first day. The change in the body weights of the henson the first day was recorded in each Experimental Section. The resultsare illustrated in the graph of FIG. 1 in the results of FIG. 1, theresults obtained by averaging the body weights of the hens included ineach Experimental Section are illustrated. Similarly, the results ofFIGS. 2 to 6 described below are the averaged values.

From the zeroth day to the seventh day, the body weights of the henswere measured and recoded in each Experimental Section and the relationbetween the number of days after the hot environment treatment startedand the body weight (g) of the hens was recorded. The results areillustrated in the graph of FIG. 2.

As is obvious from the graphs of FIG. 1 and FIG. 2, it is found that thehens in Experimental Section 1 to which the stress reliever in Example 2was fed or the hens in Experimental Section 2 to which the stressreliever in Example 1 was fed reduce the decrease in the body weightcaused by hot environment stress in the initial stage where the hotenvironment stress was applied as compared with the hens in ExperimentalSection 3 to which the bran of wheat not inoculated with the filamentousbacteria was fed. As illustrated in the graph of FIG. 2, the hens inExperimental Section 1 to which the stress reliever in Example 2containing Aspergillus oryzae that highly expressed various enzymes wasfed indicate a larger slope of the graph in the latter period of theexperiment and thus have an earlier increase in the body, weight ascompared with the hens in Experimental Section 2 to which the stressreliever in Example 1 containing Aspergillus oryzae of the wild strainwas fed.

[Observation of Change in Production Amount of Eggs]

A plurality of 259-day old hens (White Leghorn) for egg production werebred by dividing the hens from Experimental Section 5 to ExperimentalSection 7 to observe the effect in which the hot environment stressaffected change in the feedstuff ingestion amount.

[Experimental Section 5]

The feedstuff for the hens and the stress reliever in Example 2 weremixed so that the stress reliever in Example 2 was contained in thefeedstuff for the hens in 2.0% by mass and the resultant mixture was fedto the hens. As a feedstuff for the hens, the commercially availableblended feedstuff for adult chickens (High Egg, manufactured by MarubeniNisshin Feed Co., Ltd.) was used. The day when the feedstuff was changedis determined to be the zeroth day. The hens were bred in a poultryhouse where the temperature was controlled to 25° C. with an airconditioner from the zeroth day to the morning of the fourth day andbred in a poultry house having no windows under the hot environment (28°C. to 36° C.) from the afternoon of the fourth day. Experimental Section5 included seven hens.

[Experimental Section 6]

The feedstuff (High Egg, manufactured by Marubeni Nisshin Feed Co.,Ltd.) alone not including the solid-cultured product of the filamentousbacteria was fed to the hens. The hens were bred in a poultry housewhere the temperature was controlled to 25° C. with an air conditionerfrom the zeroth day to the morning of the fourth day and bred in thepoultry house having the same hot environment as the environment inExperimental Section 5 from the afternoon of the fourth day.Experimental Section 6 included five hens.

[Experimental Section 7]

Feedstuff (High Egg, manufactured by Marubeni Nisshin Feed Co., Ltd.)alone not including the solid-cultured product of the filamentousbacteria was fed to hens. The hens were bred in a poultry house wherethe temperature was controlled to 25° C. with an air conditioner fromthe zeroth day to the morning of the fourth day and bred in the poultryhouse having the same thermal neutral zone as the above saidExperimental Section 4 from the afternoon of the fourth day.Experimental Section 7 included six hens.

On the first day, the fifth day, and the eleventh day, the ingestionamounts of the feedstuff in each Experimental Section were recorded. Therelation between the number of days after the hot environment treatmentstarted and the ingestion amount (g) of the feedstuff is illustrated inthe graph of FIG. 3. The time of the ingestion of the feedstuff islimited from 15 o'clock on the day to 15 o'clock on the next day. Theingestion amount of the feedstuff was determined by weighing the feedtank before feeding and after feedstuff ingestion and calculating thedifference.

As is obvious from the graph of FIG. 3, it is found that the hens inExperimental Section 5 to which the stress reliever in Example 2 was fedhad a larger ingestion amount of the feedstuff on the eleventh day ascompared to the hens of Experimental Section 6 to which the feedstuffalone was fed. It is found that the stress reliever in Example 2 has theeffects of reducing stress, promoting recovery of the feedstuffingestion amount, and reducing the decrease in productivity when the hotenvironment stress is applied.

[Inspection of Egg Quality and the Like]

Eggs that the hens in each Experimental Section laid were collected andthe quality and the like of the eggs on the fourth day and the eleventhday were inspected. The inspection items were an egg weight, an eggshellweight, and a yolk weight. The egg weight refers to an entire weight (g)of one egg containing eggshell, egg white, and yolk. The eggshell weightrefers to the weight (g) of the eggshell per egg. The yolk weight refersto the weight (g) of yolk alone separated from the egg. The relation ofthe number of days after the hot environment treatment started and Theegg weight, eggshell weight, or yolk weight is illustrated in the graphsof FIG. 4 co FIG. 6.

As is obvious from the graphs of FIG. 4 to FIG. 6, it is found that adecrease in each of the egg weight, the eggshell weight, and the yolkweight of the hens in Experimental Section 5 to which the stressreliever in Example 2 was fed is reduced on the eleventh day as comparedto that of the hens in Experimental Section 6 to which the feedstuffalone was fed. It is found that the stress reliever in Example 2 has theeffects of reducing the decrease in the egg quality such as the eggweight, the eggshell weight, that is, the eggshell thickness, and yolkweight for a long period when the hot environment stress is applied.

As described above, feeding of the stress reliever to livestock allowsthe decrease in productivity such as the decrease in the productionamount of eggs or the deterioration in quality of eggs when the stressis applied to be reduced in the case where the livestock is livestocklaying eggs when the stress is applied to the livestock. For example,feeding of the stress reliever to livestock allows the decrease isproductivity such as the decrease in the production amount of meat orthe deterioration in quality of meat when the stress is applied to bereduced in the case where the livestock is livestock that does not layeggs. For the stress reliever, the effect of reducing a decrease is theproduction mount of leathers, hairs, or milk or deterioration in thequality of these products was also observed.

1. A method of producing a stress reliever, the method comprising:solid-culturing a substrate with filamentous bacteria to obtain asolid-cultured product, wherein the stress reliever reduces a decreasein productivity of livestock at the time of stress load.
 2. The methodof producing the stress reliever according to claim 1, wherein theproduced stress reliever reduces one or more decreases in productivityof livestock at the time of stress load by relieving stress of livestockingesting, the stress reliever, the one or more decreases being amongbody weight reduction, a decrease in an ingestion amount of feedstuff, adecrease in an egg weight, a decrease in an eggshell thickness, and adecrease in a yolk weight.
 3. The method of producing the stressreliever according to claim 1, wherein the filamentous bacteria arebacteria not producing mold poison.
 4. The method of producing thestress reliever according to claim 3, wherein the bacteria not producingmold poison are filamentous bacteria among Aspergillus oryzae,Aspergillus sojae, Aspergillus luchuensis, Aspergillus niger, orAspergillus awamori not producing mold poison.
 5. The method ofproducing the stress reliever according to claim 1, wherein thesolid-cultured product comprises polysaccharides constituting hyphae ofthe filamentous bacteria.
 6. The method of producing the stress relieveraccording to claim 1, wherein the solid-cultured product comprises anenzyme having activity.
 7. The method of producing the stress relieveraccording to claim 1, wherein the solid-cultured product comprisesviable bacteria of the filamentous bacteria.
 8. The method of producingthe stress reliever according to claim 1, further comprising:determining the solid-cultured product alone obtained by solid-culturingthe substrate with the filamentous bacteria to be the stress reliever ordetermining the solid-cultured product obtained by solid-culturing thesubstrate with the filamentous bacteria and blended with a feedingsubstance to be the stress reliever.